SUMMARY Antiretroviral therapy (ART) has significantly reduced HIV-1/AIDS-related mortality and transformed HIV infection into a chronic disease. Cardiomyopathy remains one of the leading causes of co-morbidity and heart failure in HIV-1/AIDS. However, the underlying mechanism(s) of HIV-induced cardiomyopathy and heart failure remain largely unknown. Due to a lack of HIV productive infection in cardiomyocytes, it is believed that indirect pathways involving viral and cellular factors with toxic effects are involved in HIV-induced cardiomyopathy. Among viral proteins, Nef, which is released by infected cells such as macrophages and T-cells and is taken up by neighboring cells, is a possible HIV-1 toxic factor. Nef possesses several interesting capacities that compromise multiple cellular processes including autophagy, apoptosis and viability. Our preliminary data demonstrate that Nef protein accumulates in the cardiomyocytes of HIV-infected patients and SIV-infected macaques even on ART, thus demonstrating the clinical relevance of studying Nef expression in cardiomyocytes. Accordingly, our preliminary data show that exposure of cardiomyocytes to Nef causes dysregulation of autophagy and triggers apoptosis. Mechanistically, our data indicate that Nef inhibits the terminal step of autophagy through inhibition of Beclin 1 and Rab7 localization and function. Further, we found that Nef dysregulates mitochondrial turnover, generation of ROS and decreases mitochondrial membrane potential. This leads to compromised mitochondrial function causing aberrations in cellular bioenergetic pathways. Additionally, we found that Nef reduces cellular NAD+ level, an important metabolite for cellular function and regulates cellular autophagy. Based on these observations, we hypothesize that HIV-1 Nef protein perturbs protein quality control (PQC) in cardiomyocytes by dysregulating autophagy and causes gradual cell death and consequently cardiomyopathy by impacting several critical elements that control cell homeostasis. Additionally we found that ART treatment causes mitochondrial abnormalities and inhibition of autophagy. In this application, we will examine our hypothesis by determining the role Nef and ART in cardiac PQC and cellular function in primary cardiomyocytes (Aim 1). We will investigate the molecular pathways of Nef-induced dysregulation of autophagy including the physical and functional interplay between Nef and Beclin 1, Rab7 and several other regulatory proteins involved in PQC (Aim 2). We will assess if stimulation of autophagy through NAD+ might offer a new pathway for suppressing Nef- induced pathology in cardiomyocytes (Aim 3). To address these questions, we will perform our studies in vitro using primary cardiomyocytes, validate in animal models (HIV-1 TG26 and transgenic Nef) and confirm in HIV+ clinical samples with ART. The outcome of these studies will unravel the molecular mechanism involved in HIV- induced cardiomyopathy and will help to develop intelligent strategies to prevent HIV-1 associated cardiomyopathy.